title: Activation of extracellular signal-regulated kinase 1/2 and Sp1 may contribute to the expression of tissue inhibitor of metalloproteinases-1 induced by transforming growth factor-β1 in human pulmonary arterial smooth muscle cells.
abstract: OBJECTIVE
  Tissue inhibitor of metalloproteinases-1 (TIMP-1) plays an important role in the development of pulmonary arterial hypertension. However, the molecular regulatory mechanisms of TIMP-1 in the pulmonary arteries are not fully understood, especially in human pulmonary arterial smooth muscle cells (HPASMCs). We investigated the signaling pathway involved in the regulation of TIMP-1 in HPASMCs induced by transforming growth factor (TGF)-β1.
  
  
  METHODS
  Cultured HPASMCs were incubated with different concentrations of TGF-β1 (0-40 ng/mL) for 24 h or with 10 ng/mL TGF-β1 for different times (1-48 h).
  
  
  RESULTS
  Western blot, real-time polymerase chain reaction and enzyme-linked immunosorbent assay analyses showed that TGF-β1 enhanced the expression and secretion of TIMP-1 in a time-dependent and dose-dependent fashion. TGF-β1 could phosphorylate two of the three mitogen-activated protein kinases-extracellular signal-regulated kinase 1/2 (ERK1/2) and p38, but not c-Jun NH2-terminal kinase. Of these kinases, only the inhibition of ERK1/2 by U0126, which was a specific inhibitor of mitogen-activated protein kinase/ERK1/2, effectively blocked the TGF-β1-induced expression of TIMP-1. Mithramycin, an inhibitor of Sp1 transcription factor, also significantly inhibited the expression of TIMP-1. Additionally, electrophoretic mobility shift assay showed that TGF-β1 could up-regulate the DNA-binding activity of Sp1 and that U0126 and mithramycin could effectively inhibit these events.
  
  
  CONCLUSIONS
  TGF-β1 could stimulate the expression and secretion of TIMP-1 in HPASMCs in a time-dependent and dose-dependent fashion, and ERK1/2 and Sp1 signaling pathways might be involved in these activities.
authors.affiliation: Department of Cardiology, Shaoxing People's Hospital (Shaoxing Hospital of Zhejiang University), Shaoxing City, Zhejiang, PR China; Department of Cardiology, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou City, Zhejiang, PR China.
authors.forename: Weiliang
authors.initials: W
authors.lastname: Tang
authors.affiliation: Department of Cardiology, Zhejiang Traditional Chinese Medical Hospital, Hangzhou City, Zhejiang, PR China.
authors.forename: Jinxiu
authors.initials: J
authors.lastname: Yang
authors.affiliation: Department of Cardiology, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou City, Zhejiang, PR China.
authors.forename: Furong
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Department of Cardiology, Shaoxing People's Hospital (Shaoxing Hospital of Zhejiang University), Shaoxing City, Zhejiang, PR China.
authors.forename: Hangyuan
authors.initials: H
authors.lastname: Guo
authors.affiliation: Department of Cardiology, Shaoxing People's Hospital (Shaoxing Hospital of Zhejiang University), Shaoxing City, Zhejiang, PR China.
authors.forename: Fang
authors.initials: F
authors.lastname: Peng
authors.affiliation: Department of Cardiology, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou City, Zhejiang, PR China. Electronic address: Wangxx0571@yahoo.com.cn.
authors.forename: Xingxiang
authors.initials: X
authors.lastname: Wang
mesh_terms: D002070:Butadienes / Q000494:pharmacology; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D006976:Hypertension, Pulmonary / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D009570:Nitriles / Q000494:pharmacology; D008926:Plicamycin / Q000494:pharmacology; D011651:Pulmonary Artery / Q000473:pathology*; D016329:Sp1 Transcription Factor / Q000037:antagonists & inhibitors; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*
chemical_list: D002070:Butadienes; D009570:Nitriles; D016329:Sp1 Transcription Factor; C560822:TIMP1 protein, human; D019715:Tissue Inhibitor of Metalloproteinase-1; D053773:Transforming Growth Factor beta1; C113580:U 0126; D019950:Mitogen-Activated Protein Kinase 1; D008926:Plicamycin
keywords: Sp1; extracellular signal-regulated kinase 1/2; human pulmonary arterial smooth muscle cells; tissue inhibitor of metalloproteinase-1; transforming growth factor-β1